This research is designed to determine the extent to which the adverse effects of the transfusion of stored RBCs result from the acute delivery of iron to the monocyte-macrophage system. Although transfusion is effective in treatment of symptomatic anemia, there is increasing evidence that RBC transfusions increase rates of infection, morbidity, and mortality in hospitalized patients. Our preliminary data in mice and humans strongly suggest that this is indeed the case, and that iron is the most likely culprit in producing these adverse effects. By current FDA standards, a unit of stored red blood cells (RBC) is clinically acceptable for transfusion if as much as 25% of the RBC are cleared within 24 hours, thereby delivering a substantial dose of iron to the monocyte-macrophage system. The central hypothesis of this proposal is that iron, acting both intracellularly in phagocytes and extracellularly as plasma non-transferrin bound iron (NTBI), is responsible for the adverse effects associated with stored RBC transfusions. To accomplish our aims, we will use well-characterized mouse models of transfusion and sepsis. This research has three Specific Aims: Aim #1: To test the hypothesis that the pro-oxidant effects of increased intracellular iron levels are responsible for the observed pro-inflammatory state following transfusion of older, stored RBCs. Aim #2: To test the hypothesis that the acute clearance of older stored RBCs alters T helper (TH) cell differentiation towards TH17 cells. Aim #3: To test the hypothesis that NTBI released following the acute clearance of older stored RBCs increases the severity of infection in models of monobacterial infection and polymicrobial sepsis. Thus, this proposal will define the mechanism of the iron insult in Aim #1, characterize the immune response to this insult in Aim #2, and study the clinical consequences of this insult in Aim #3. The insights gained from this project will guide future pre-clinical studies evaluating novel therapeutic approaches in human transfusion medicine. This project, along with a well-developed career development plan, including four distinguished mentors and advisors, will provide the applicant with a training vehicle in immunology and iron biology, which will advance his research career to that of an independent investigator.